Animal-derived surfactant preparations are highly effective in preventing and treating respiratory distress syndrome (RDS) in premature infants, but not in pediatric and adult patients with acute lung injury (ALI) and the acute respiratory distress syndrome (ARDS). Effective therapy for ALI/ARDS will probably require synthetic surfactant with maximal surface activity and the ability to resist surfactant inhibition due to vascular leakage and acute inflammation. Such synthetic preparations offer the advantages of large-scale production, compositional reproducibility, easier quality control, stability with a long shelf-life, no transmission of animal disease, and lower costs. Synthetic surfactants also allow the use of novel protein and lipid components that may be more inhibition-resistant and possess biophysical properties that outperform native surfactant formulations than are presently available. Our laboratory has designed, synthesized, and characterized several highly-active analogs of surfactant protein B (SP-B), such as "Mini-B" (i.e., cross-linked construct of the N- and C-terminal domains of SP-B) and "Super Mini-B" (i.e., Mini-B with an insertion sequence at the N-terminus), and observed that Super-Mini-B further enhances the surface activity of synthetic surfactant preparations over that seen with native SP-B proteins. Mixing these SP-B analogs in non-toxic phospholipase-resistant lipids, i.e. phosphonolipid analogs of dipalmitoyl phosphatidylcholine (DPPC) and phosphatidylglycerol (PG), will promote inhibition resistance. The specific aims of this proposal are to (1) re-engineer, synthesize and characterize novel synthetic SP-B analogs designed to have high lipid-binding, surface activity and resistance to inhibition and oxidative stress in (phospholipase-resistant) lipid mixtures, and (2) define the in vivo efficacy of synthetic surfactants in rabbits with oleic acid- or lipopolysaccharide (LPS)-induced acute lung injury. We hypothesize that additional re-engineering of Mini-B and Super Mini-B, based on using discrete amino acid substitutions, will maximize their respective lipid binding and surface activity, and also increase their resistance to inhibition and oxidative stress. Formulation of these re-engineered SP-B analogs in phospholipase-resistant lipid mixtures may deliver a synthetic surfactant preparation uniquely able to treat ALI/ARDS. PROJECT NARRATIVE. Acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) are life-threatening respiratory diseases, which affect children and adults with sepsis, pneumonia, gastric aspiration, burns, trauma, and other acute illnesses. Survival depends on assisted ventilation, treatment of the underlying disease, and reversal of surfactant inhibition in the lung. Re-engineering of the essential surfactant protein B and the predominant lipids to enhance resistance to inhibition and oxidative stress may deliver a synthetic surfactant preparation uniquely able to treat ALI/ARDS.